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Glioblastoma (GBM) is a poorly treatable disease due to the fast development of tumor recurrences and high resistance to chemo- and radiotherapy. To overcome the highly adaptive behavior of GBMs, especially multimodal therapeutic approaches also including natural adjuvants have been investigated. However, despite increased efficiency, some GBM cells are still able to survive these advanced treatment regimens. Given this, the present study evaluates representative chemoresistance mechanisms of surviving human GBM primary cells in a complex in vitro co-culture model upon sequential application of temozolomide (TMZ) combined with AT101, the R(-) enantiomer of the naturally occurring cottonseed-derived gossypol. Treatment with TMZ+AT101/AT101, although highly efficient, yielded a predominance of phosphatidylserine-positive GBM cells over time. Analysis of the intracellular effects revealed phosphorylation of AKT, mTOR, and GSK3ß, resulting in the induction of various pro-tumorigenic genes in surviving GBM cells. A Torin2-mediated mTOR inhibition combined with TMZ+AT101/AT101 partly counteracted the observed TMZ+AT101/AT101-associated effects. Interestingly, treatment with TMZ+AT101/AT101 concomitantly changed the amount and composition of extracellular vesicles released from surviving GBM cells. Taken together, our analyses revealed that even when chemotherapeutic agents with different effector mechanisms are combined, a variety of chemoresistance mechanisms of surviving GBM cells must be taken into account.
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Neoplasias Encefálicas , Glioblastoma , Gosipol , Humanos , Temozolomida/farmacología , Temozolomida/uso terapéutico , Glioblastoma/tratamiento farmacológico , Glioblastoma/genética , Gosipol/farmacología , Resistencia a Antineoplásicos/genética , Línea Celular Tumoral , Recurrencia Local de Neoplasia/tratamiento farmacológico , Serina-Treonina Quinasas TOR , Neoplasias Encefálicas/tratamiento farmacológico , Neoplasias Encefálicas/genética , Antineoplásicos Alquilantes/farmacología , Antineoplásicos Alquilantes/uso terapéuticoRESUMEN
Gossypol, a sesquiterpenoid found in cotton seeds, exerts anticancer effects on several tumor entities due to inhibition of DNA synthesis and other mechanisms. In clinical oncology, histone deacetylase inhibitors (HDACi) are applied as anticancer compounds. In this study, we examined whether gossypol harbors HDAC inhibiting activity. In vitro analyses showed that gossypol inhibited class I, II, and IV HDAC, displaying the capability to laterally interact with the respective catalytic center and is, therefore, classified as a pan-HDAC inhibitor. Next, we studied the effects of gossypol on human-derived hepatoma (HepG2) and colon carcinoma (HCT-116) cell lines and found that gossypol induced hyperacetylation of histone protein H3 and/or tubulin within 6 h. Furthermore, incubation with different concentrations of gossypol (5-50 µM) over a time period of 96 h led to a prominent reduction in cellular viability and proliferation of hepatoma (HepG2, Hep3B) and colon carcinoma (HCT-116, HT-29) cells. In-depth analysis of underlying mechanisms showed that gossypol induced apoptosis via caspase activation. For pre-clinical evaluation, toxicity analyses showed toxic effects of gossypol in vitro toward non-malignant primary hepatocytes (PHH), the colon-derived fibroblast cell line CCD-18Co, and the intestinal epithelial cell line CCD 841 CoN at concentrations of ≥5 µM, and embryotoxicity in chicken embryos at ≥2.5 µM. In conclusion, the pronounced inhibitory capacity of gossypol on cancer cells was characterized, and pan-HDACi activity was detected in silico, in vitro, by inhibiting individual HDAC isoenzymes, and on protein level by determining histone acetylation. However, for clinical application, further chemical optimization is required to decrease cellular toxicity.
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The potential of gossypol and of its R-(-)-enantiomer (R-(-)-gossypol acetic acid, AT-101), has been evaluated for treatment of cancer as an independent agent and in combination with standard chemo-radiation-therapies, respectively. This review assesses the evidence for safety and clinical effectiveness of oral gossypol/AT-101 in treating various types of cancer. The databases PubMed, MEDLINE, Cochrane, and ClinicalTrials.gov were examined. Phase I and II trials as well as single arm and randomized trials were included in this review. Results were screened to determine if they met inclusion criteria and then summarized using a narrative approach. A total of 17 trials involving 759 patients met the inclusion criteria. Overall, orally applied gossypol/AT-101 at low doses (30 mg daily or lower) was determined as well tolerable either as monotherapy or in combination with chemo-radiation. Adverse events should be strictly monitored and were successfully managed by dose-reduction or treating symptoms. There are four randomized trials, two performed in patients with advanced non-small cell lung cancer, one in subjects with head and neck cancer, and one in patients with metastatic castration-resistant prostate cancer. Thereby, standard chemotherapy (either docetaxel (two trials) or docetaxel plus cisplatin or docetaxel plus prednisone) was tested with and without AT-101. Within these trials, a potential benefit was observed in high-risk patients or in some patients with prolongation in progression-free survival or in overall survival. Strikingly, the most recent clinical trial combined low dose AT-101 with docetaxel, fluorouracil, and radiation, achieving complete responses in 11 of 13 patients with gastroesophageal carcinoma (median duration of 12 months) and a median progression-free survival of 52 months. The promising results shown in subsets of patients supports the need of further specification of AT-101 sensitive cancers as well as for the establishment of effective AT-101-based therapy. In addition, the lowest recommended dose of gossypol and its precise toxicity profile need to be confirmed in further studies. Randomized placebo-controlled trials should be performed to validate these data in large cohorts.
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INTRODUCTION: (-)-Gossypol (AT-101), the (-)-enantiomer of the natural compound gossypol, has shown significant inhibitory effects on various types of cancers such as osteosarcoma, myeloma, glioma, lung cancer, and prostate cancer. However, the clinical application of (-)-gossypol was often hindered by its evident side effects and the low bioavailability via oral administration, which necessitated the development of suitable (-)-gossypol preparations to settle the problems. In this study, injectable cyclic RGD (cRGD)-decorated liposome (cRGD-LP) was prepared for tumor-targeted delivery of (-)-gossypol. METHODS: The cRGD-LP was prepared based on cRGD-modified lipids. For comparison, a non-cRGD-containing liposome (LP) with a similar chemical composition to cRGD-LP was specially designed. The physicochemical properties of (-)-gossypol-loaded cRGD-LP (Gos/cRGD-LP) were investigated in terms of the drug loading efficiency, particle size, morphology, drug release, and so on. The inhibitory effect of Gos/cRGD-LP on the proliferation of tumor cells in vitro was evaluated using different cell lines. The biodistribution of cRGD-LP in vivo was investigated via the near-infrared (NIR) fluorescence imaging technique. The antitumor effect of Gos/cRGD-LP in vivo was evaluated in PC-3 tumor-bearing nude mice. RESULTS: Gos/cRGD-LP had an average particle size of about 62 nm with a narrow size distribution, drug loading efficiency of over 90%, and sustained drug release for over 96 h. The results of NIR fluorescence imaging demonstrated the enhanced tumor targeting of cRGD-LP in vivo. Moreover, Gos/cRGD-LP showed a significantly enhanced inhibitory effect on PC-3 tumors in mice, with a tumor inhibition rate of over 74% and good biocompatibility. CONCLUSION: The incorporation of cRGD could significantly enhance the tumor-targeting effect of the liposomes and improve the antitumor effect of the liposomal (-)-gossypol in vivo, which indicated the potential of Gos/cRGD-LP that warrants further investigation for clinical applications of this single-isomer drug.
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Gosipol , Liposomas , Animales , Línea Celular Tumoral , Gosipol/análogos & derivados , Masculino , Ratones , Ratones Desnudos , Péptidos Cíclicos , Distribución TisularRESUMEN
The aim of this study was to determine the appropriate doses of AT-101 and resveratrol combination in the in vitro hormone-refractory prostate cancer (PC) cell lines, in order to evaluate the cytotoxic and genotoxic effects of this combination on the proliferation of cancer cells, namely PC-3, DU-145 and LNCAP. Cytotoxicity in PC cell lines was analysed by using the XTT Cell Proliferation Assay. DNA damage was performed with the cell death assay. Apoptotic protein levels were performed by Roche Human Apoptosis Array. IC50 values were determined by XTT analysis. The strongest combined doses (100 µM resveratrol + 5µM AT-101) were found to have the strongest synergistic apoptotic and cytotoxic effects on DU-145 cells at 72 hr. While the combined use of resveratrol and AT-101 increased the expression of markers in apoptotic cell pathways on cells, a decrease in the expression of anti-apoptotic markers was detected (p Ë 0.05). Combined applications of these compounds showed an important synergism in the hormone-refractory PC cell lines, and it was determined that after the post-translational modification, they were significantly effective on the apoptotic pathway. These results have revealed that the combination of resveratrol and AT-101 holds great expectation as a new chemotherapeutic application in the treatment of human prostate cancer.
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Apoptosis , Neoplasias de la Próstata , Línea Celular , Línea Celular Tumoral , Gosipol/análogos & derivados , Hormonas , Humanos , Masculino , Neoplasias de la Próstata/tratamiento farmacológico , Resveratrol/farmacologíaRESUMEN
OBJECTIVES: The Inhibitors of Apoptosis (IAPs) regulate initiator and effector phases of caspase mediated apoptosis. This study evaluates the effects of SMAC mimetic AT-101 in regulation of IAPs/caspases/NFÆB-p65 in an adenocarcinoma cell line. MATERIALS AND METHODS: MTT assay was performed in the NCI-H522 cell line. Flow cytometry was used for detecting cell cycle, apoptosis, and NFÆB-p65 regulation. Effects of AT-101 on IAPs and caspases were determined by quantitative real time-PCR and western blotting. AutoDock-VINA was used for computational analysis. RESULTS: AT-101 reduced the cell proliferation of NCI-H522 with a GI50 value of 7 µM. The compound arrested adenocarcinoma cells in the G1 phase of the cell cycle and increased early and late phase apoptosis while decreasing tumor-cell trans-migration. AT-101 treatment to NCI H522 at a concentration of 0.35 µM decreased XIAP, cIAP-1, and cIAP-2 mRNA levels to 4.39±0.66, 1.93±0.26, and 2.20±0.24 folds, respectively. Increased dose of AT-101 at 0.7 µM concentration further decreased XIAP, cIAP-1, and cIAP-2 mRNA levels to 2.44±0.67, 1.46±0.93, and 0.97±0.10 folds, respectively. Similar effects of a dose-dependent decrease in the protein expressions of XIAP, cIAP-1, and cIAP-2 were observed with AT-101 treatments, while a dose-responsive increase in the mRNA and protein expression levels of caspase 6 and caspase 7 was observed in the NCI-H522 cell line. The compound exhibited binding affinity (-6.1 kcal/mol) and inhibited NFÆB-p65 in these cells. CONCLUSION: AT-101 had anti-tumor efficacy against lung adenocarcinoma cells which could be mediated through IAPs/caspase-dependent apoptosis and NFÆB-p65 cross talk. Results from this study suggests a signal cross talk between IAPs and NFkB and open new channels for further investigations in therapeutic intervention against lung cancer management.
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(1) Purpose: The objective was to evaluate CT perfusion and radiomic features for prediction of one year disease free survival in laryngeal and hypopharyngeal cancer. (2) Method and Materials: This retrospective study included pre and post therapy CT neck studies in 36 patients with laryngeal/hypopharyngeal cancer. Tumor contouring was performed semi-autonomously by the computer and manually by two radiologists. Twenty-six radiomic features including morphological and gray-level features were extracted by an internally developed and validated computer-aided image analysis system. The five perfusion features analyzed included permeability surface area product (PS), blood flow (flow), blood volume (BV), mean transit time (MTT), and time-to-maximum (Tmax). One year persistent/recurrent disease data were obtained following the final treatment of definitive chemoradiation or after total laryngectomy. We performed a two-loop leave-one-out feature selection and linear discriminant analysis classifier with generation of receiver operating characteristic (ROC) curves and confidence intervals (CI). (3) Results: 10 patients (28%) had recurrence/persistent disease at 1 year. For prediction, the change in blood flow demonstrated a training AUC of 0.68 (CI 0.47-0.85) and testing AUC of 0.66 (CI 0.47-0.85). The best features selected were a combination of perfusion and radiomic features including blood flow and computer-estimated percent volume changes-training AUC of 0.68 (CI 0.5-0.85) and testing AUC of 0.69 (CI 0.5-0.85). The laryngoscopic percent change in volume was a poor predictor with a testing AUC of 0.4 (CI 0.16-0.57). (4) Conclusions: A combination of CT perfusion and radiomic features are potential predictors of one-year disease free survival in laryngeal and hypopharyngeal cancer patients.
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Neoplasias Hipofaríngeas , Supervivencia sin Enfermedad , Humanos , Neoplasias Hipofaríngeas/diagnóstico por imagen , Neoplasias Hipofaríngeas/cirugía , Recurrencia Local de Neoplasia , Perfusión , Proyectos Piloto , Estudios Retrospectivos , Tomografía Computarizada por Rayos XRESUMEN
Human aspartate/asparagine-ß-hydroxylase (AspH) is a 2-oxoglutarate (2OG) dependent oxygenase that catalyses the hydroxylation of Asp/Asn-residues of epidermal growth factor-like domains (EGFDs). AspH is reported to be upregulated on the cell surface of invasive cancer cells in a manner distinguishing healthy from cancer cells. We report studies on the effect of small-molecule active pharmaceutical ingredients (APIs) of human cancer therapeutics on the catalytic activity of AspH using a high-throughput mass spectrometry (MS)-based inhibition assay. Human B-cell lymphoma-2 (Bcl-2)-protein inhibitors, including the (R)-enantiomer of the natural product gossypol, were observed to efficiently inhibit AspH, as does the antitumor antibiotic bleomycin A2. The results may help in the design of AspH inhibitors with the potential of increased selectivity compared to the previously identified Fe(II)-chelating or 2OG-competitive inhibitors. With regard to the clinical use of bleomycin A2 and of the Bcl-2 inhibitor venetoclax, the results suggest that possible side-effects mediated through the inhibition of AspH and other 2OG oxygenases should be considered.
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Antibióticos Antineoplásicos/farmacología , Bleomicina/farmacología , Inhibidores Enzimáticos/farmacología , Gosipol/farmacología , Oxigenasas de Función Mixta/antagonistas & inhibidores , Bibliotecas de Moléculas Pequeñas/farmacología , Antibióticos Antineoplásicos/química , Bleomicina/química , Relación Dosis-Respuesta a Droga , Composición de Medicamentos , Inhibidores Enzimáticos/química , Gosipol/química , Humanos , Oxigenasas de Función Mixta/aislamiento & purificación , Oxigenasas de Función Mixta/metabolismo , Simulación del Acoplamiento Molecular , Estructura Molecular , Bibliotecas de Moléculas Pequeñas/química , Relación Estructura-ActividadRESUMEN
Anti-apoptotic BCL2 proteins are important targets for cancer therapy as cancers depend on their activity for survival. Direct inhibitors of MCL1 have entered clinical trials, although their efficacy may be limited by toxicity. An alternative approach may be to induce the pro-apoptotic protein NOXA which selectively inhibits MCL1 in cells. Many compounds originally proposed as inhibitors of the BCL2 family were subsequently found to induce the pro-apoptotic protein NOXA through the unfolded protein response. In the present study, we compared various putative BH3 mimetics across a panel of carcinoma cell lines and measured expression of NOXA protein and mRNA, as well as the kinetics of NOXA induction. We found that AT101 [(-)-gossypol] induces high levels of NOXA in carcinoma cell lines yet cells survive. When combined with an appropriate BCL2 or BCL-XL inhibitor, NOXA-dependent sensitization occurs. NOXA protein continues to accumulate for many hours after AT101 is removed, providing a window for administering these combinations. As MCL1 promotes drug resistance and overall survival, we propose that NOXA induction is an alternative therapeutic strategy to target MCL1 and either kill cancer cells that are dependent on MCL1 or sensitize cancer cells to other BCL2 inhibitors.
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Aim: AT-101 is a polyphenolic compound with potent anti-apoptotic effects in various cancers. In this study, the possible synergistic cytotoxic and apoptotic effect of trastuzumab/AT-101 combination was investigated in HER2-positive breast cancer cell lines. Materials & methods: SKBR-3, MDA-MB-453 and MCF-10A cell lines were treated with a trastuzumab/AT-101 combination. Synergistic cytotoxicity and apoptosis effects were shown and then PI3K and Akt protein levels were studied. Result: The trastuzumab/AT-101 combination induced synergistic cytotoxicity and apoptosis in both breast cancer cells but not in MCF-10A cells. Combination treatment induced cytotoxicity via inhibiting PI3K/AKT but not the MAPK/ERK pathway. Conclusion: The trastuzumab/AT-101 combination may be a good candidate for patients with trastuzumab-resistant Her2-positive breast cancer and inhibition of the PI3K/AKT pathway may be one of the underlying mechanisms.
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Protocolos de Quimioterapia Combinada Antineoplásica/farmacología , Neoplasias de la Mama/tratamiento farmacológico , Gosipol/análogos & derivados , Trastuzumab/farmacología , Antineoplásicos Inmunológicos/farmacología , Antineoplásicos Inmunológicos/uso terapéutico , Antineoplásicos Fitogénicos/farmacología , Antineoplásicos Fitogénicos/uso terapéutico , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Apoptosis/efectos de los fármacos , Neoplasias de la Mama/patología , Línea Celular Tumoral , Ensayos de Selección de Medicamentos Antitumorales , Sinergismo Farmacológico , Femenino , Gosipol/farmacología , Gosipol/uso terapéutico , Humanos , Fosfatidilinositol 3-Quinasas/metabolismo , Proteínas Proto-Oncogénicas c-akt/metabolismo , Receptor ErbB-2/metabolismo , Transducción de Señal/efectos de los fármacos , Trastuzumab/uso terapéuticoRESUMEN
BACKGROUND: The treatment of glioblastomas (GBM) is still a clinical challenge. Current GBM therapeutic plans focus on the development of new strategies for local drug administration in the tumor cavity to realize an efficient long-term treatment with small side-effects. Here, different amounts of residual GBM cells and healthy brain cells define the microenvironment of the tumor cavity after individual surgical GBM resection (complete or incomplete). METHODS: We evaluated available in vivo data and determined the required amounts and numerical ratios of GBM and healthy brain cells for our in vitro (in)complete resection dual co-culture model. We applied a generic two-drug treatment [Temozolomide (TMZ) in combination with AT101, followed by single AT101 treatment] strategy and analyzed the results in comparison with appropriate mono-culture systems to prove the applicability of our model. RESULTS: We established a suitable GBM dual co-culture model, mimicking the complete and incomplete resection in vitro, giving stable and reliable results on drug testing. Both dual co-culture conditions protectively influenced on cell death and growth rates of primary GBMs when treated with TMZ+AT101/AT101, although the treatment strategy per se was still efficient. Cell death of astrocytes correlated with amounts of increasing GBM cell numbers in the incomplete resection model upon drug treatment, and probably GBM-released chemokine and cytokines were involved in this interplay. CONCLUSIONS: Our results suggest that this dual co-culture model provides a biologically relevant platform for the discovery and compound screening of local GBM treatment strategies.
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Antineoplásicos Alquilantes/toxicidad , Antineoplásicos Fitogénicos/toxicidad , Astrocitos/citología , Glioblastoma/patología , Microglía/citología , Análisis de Varianza , Astrocitos/efectos de los fármacos , Encéfalo/citología , Técnicas de Cocultivo , Glioblastoma/tratamiento farmacológico , Glioblastoma/cirugía , Gosipol/análogos & derivados , Gosipol/toxicidad , Humanos , Microglía/efectos de los fármacos , Temozolomida/toxicidadRESUMEN
PURPOSE: Glioblastoma multiforme (GBM) is a poorly curable disease due to its profound chemoresistance. Despite recent advances in surgery, radiotherapy and chemotherapy, the efficient treatment of GBMs is still a clinical challenge. Beside others, AT101, the R-(-) enantiomer of gossypol, and demethoxycurcumin (DMC), a curcumin-related demethoxy compound derived from Curcuma longa, were considered as possible alternative drugs for GBM therapy. METHODS: Using different human primary GBM cell cultures in a long-term stimulation in vitro model, the cytotoxic and anti-proliferative effects of single and combined treatment with 5 µM AT101 and 5 µM or 10 µM DMC were investigated. Furthermore, western blots on pAkt and pp44/42 as well as JC-1 staining and real-time RT-PCR were performed to understand the influence of the treatment at the molecular and gene level. RESULTS: Due to enhanced anti-proliferative effects, we showed that combined therapy with both drugs was superior to a single treatment with AT101 or DMC. Here, by determination of the combination index, a synergism of the combined drugs was detectable. Phosphorylation and thereby activation of the kinases p44/42 and Akt, which are involved in proliferation and survival processes, were inhibited, the mitochondrial membrane potential of the GBM cells was altered, and genes involved in dormancy-associated processes were regulated by the combined treatment strategy. CONCLUSION: Combined treatment with different drugs might be an option to efficiently overcome chemoresistance of GBM cells in a long-term treatment strategy.
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Protocolos de Quimioterapia Combinada Antineoplásica/farmacología , Neoplasias Encefálicas/tratamiento farmacológico , Diarilheptanoides/farmacología , Glioblastoma/tratamiento farmacológico , Gosipol/análogos & derivados , Antineoplásicos Fitogénicos/administración & dosificación , Antineoplásicos Fitogénicos/farmacología , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/patología , Proliferación Celular/efectos de los fármacos , Diarilheptanoides/administración & dosificación , Relación Dosis-Respuesta a Droga , Sinergismo Farmacológico , Expresión Génica/efectos de los fármacos , Glioblastoma/genética , Glioblastoma/metabolismo , Glioblastoma/patología , Gosipol/administración & dosificación , Gosipol/farmacología , Humanos , Potencial de la Membrana Mitocondrial/efectos de los fármacos , Transducción de Señal/efectos de los fármacos , Células Tumorales CultivadasRESUMEN
Osteosarcoma (OS) is the most common primary malignant bone tumor and mainly affects children and adolescents. The OS five-year survival rate remains very low. Thus, novel therapeutic protocols for the treatment of OS are needed. Several approaches targeting deregulated signaling pathways have been proposed. The antitumoral effects of polyphenols, which are naturally occurring compounds with potent antioxidant and anti-inflammatory activity, have been investigated in different tumors. Gossypol, which is a natural polyphenolic aldehyde isolated from the seeds of the cotton plant, has been shown to exert antitumoral activity in leukemia and lymphoma and in breast, head and neck, colon and prostate cancers. Therefore, in this study, we evaluated the effect of AT-101, which is the (-) enantiomer and more active form of gossypol, on the growth of human and murine OS cells in vitro and in vivo. Several clinical trials employing AT-101 have been performed, and some clinical trials are ongoing. Our results showed for the first time that AT-101 significantly inhibits OS cell growth in a dose- and time-dependent manner, inducing apoptosis and necrosis and partially activating autophagy. Our results demonstrated that AT-101 inhibits prosurvival signaling pathways depending on Akt, p38 MAPK and JNK. In addition, treatment with AT-101 increases the survival of OS-bearing mice. Overall, these results suggest that AT-101 is a candidate chemo-supportive molecule for the development of novel chemotherapeutic protocols for the treatment of OS.
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Antineoplásicos/farmacología , Neoplasias Óseas/tratamiento farmacológico , Gosipol/análogos & derivados , Osteosarcoma/tratamiento farmacológico , Proteínas Proto-Oncogénicas c-bcl-2/antagonistas & inhibidores , Animales , Apoptosis/efectos de los fármacos , Autofagia/efectos de los fármacos , Neoplasias Óseas/metabolismo , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Gosipol/farmacología , Humanos , Sistema de Señalización de MAP Quinasas/efectos de los fármacos , Ratones , Ratones Endogámicos BALB C , Osteosarcoma/metabolismo , Polifenoles/farmacología , Especies Reactivas de Oxígeno/metabolismo , Transducción de Señal/efectos de los fármacos , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismoRESUMEN
Autophagy has important functions in maintaining energy metabolism under conditions of starvation and to alleviate stress by removal of damaged and potentially harmful cellular components. Therefore, autophagy represents a pro-survival stress response in the majority of cases. However, the role of autophagy in cell survival and cell death decisions is highly dependent on its extent, duration, and on the respective cellular context. An alternative pro-death function of autophagy has been consistently observed in different settings, in particular, in developmental cell death of lower organisms and in drug-induced cancer cell death. This cell death is referred to as autophagic cell death (ACD) or autophagy-dependent cell death (ADCD), a type of cellular demise that may act as a backup cell death program in apoptosis-deficient tumors. This pro-death function of autophagy may be exerted either via non-selective bulk autophagy or excessive (lethal) removal of mitochondria via selective mitophagy, opening new avenues for the therapeutic exploitation of autophagy/mitophagy in cancer treatment.
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Leukemia stem cells (LSCs) are deemed to the mainspring for treatment failure in acute myeloid leukemia (AML). Conventional chemotherapeutic drugs fail to eradicate leukemia stem cells, which becomes the root of drug resistance and disease recurrence. Hence, new therapeutic strategies targeting LSCs are supposed to be critical for patients with AML. Here we report that combination of Bcl-2 inhibitor AT-101 and chemotherapeutic drug idarubicin (IDA) results in synergistic lethality in CD34+CD38- leukemia stem-like cells sorted from KG-1α and Kasumi-1 AML cell lines and primary CD34+ AML cells in vitro while sparing the normal counterparts. In addition, combinatorial treatment also significantly inhibits the growth of patient-derived xenograft (PDX) mouse models generated from FLT3-ITDmut AML patient in vivo. Mechanistically, the synergistic effects of AT-101 with IDA to induce cell death are closely associated with blockage of DNA damage repair and thus activates the intrinsic apoptotic pathway. In summary, these findings suggest that combinatorial therapy with AT-101 and IDA selectively eliminates leukemia stem-like cells both in vitro and in vivo, representing a potent and alternative salvage therapy for the treatment of relapsed and refractory patients with AML.
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Protocolos de Quimioterapia Combinada Antineoplásica/farmacología , Apoptosis/efectos de los fármacos , Reparación del ADN/efectos de los fármacos , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Leucemia Mieloide Aguda/tratamiento farmacológico , Leucemia Mieloide Aguda/patología , Mutaciones Letales Sintéticas/efectos de los fármacos , Adolescente , Adulto , Anciano , Animales , Biomarcadores de Tumor/genética , Estudios de Casos y Controles , Proliferación Celular/efectos de los fármacos , Reparación del ADN/genética , Sinergismo Farmacológico , Femenino , Estudios de Seguimiento , Gosipol/administración & dosificación , Gosipol/análogos & derivados , Humanos , Idarrubicina/administración & dosificación , Leucemia Mieloide Aguda/genética , Masculino , Ratones , Ratones Endogámicos NOD , Ratones SCID , Persona de Mediana Edad , Pronóstico , Proteína 1 Compañera de Translocación de RUNX1/antagonistas & inhibidores , Proteína 1 Compañera de Translocación de RUNX1/genética , Tasa de Supervivencia , Células Tumorales Cultivadas , Ensayos Antitumor por Modelo de Xenoinjerto , Adulto Joven , Tirosina Quinasa 3 Similar a fms/antagonistas & inhibidores , Tirosina Quinasa 3 Similar a fms/genéticaRESUMEN
Background Adrenal cortical carcinoma (ACC) is a rare cancer with treatment options of limited efficacy, and poor prognosis if metastatic. AT-101 is a more potent inhibitor of B cell lymphoma 2 family apoptosis-related proteins than its racemic form, gossypol, which showed preliminary clinical activity in ACC. We thus evaluated the efficacy of AT-101 in patients with advanced ACC. Methods Patients with histologically confirmed metastatic, recurrent, or primarily unresectable ACC were treated with AT-101 (20 mg/day orally, 21 days out of 28-day cycles) until disease progression and/or prohibitive toxicity. The primary endpoint was objective response rate, wherein a Response Evaluation Criteria In Solid Tumors (RECIST) partial response rate of 25% would be considered promising and 10% not, with a Type I error of 10% and 90% power. In a 2-stage design, 2 responses were required of the first 21 assessable subjects to warrant complete accrual of 44 patients. Secondary endpoints included safety, progression-free survival and overall survival. Results This study accrued 29 patients between 2009 and 2011; median number of cycles was 2. Seven percent experienced grade 4 toxicity including cardiac troponin elevations and hypokalemia. None of the first 21 patients attained RECIST partial response; accordingly, study therapy was deemed ineffective and the trial was permanently closed. Conclusions AT-101 had no meaningful clinical activity in this study in patients with advanced ACC, but demonstrated feasibility of prospective therapeutic clinical trials in this rare cancer.
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Neoplasias de la Corteza Suprarrenal/tratamiento farmacológico , Carcinoma Corticosuprarrenal/tratamiento farmacológico , Antineoplásicos Fitogénicos/uso terapéutico , Gosipol/análogos & derivados , Adulto , Anciano , Antineoplásicos Fitogénicos/efectos adversos , Femenino , Gosipol/efectos adversos , Gosipol/uso terapéutico , Humanos , Masculino , Persona de Mediana Edad , Proteínas Proto-Oncogénicas c-bcl-2 , Criterios de Evaluación de Respuesta en Tumores SólidosRESUMEN
Malignant mesothelioma (MM) is a primary tumor arising from mesothelial cells. The survival of MM patients following traditional chemotherapy is poor, thus innovative treatments for MM are needed. (-)-gossypol (AT-101) is a BH3 mimetic compound which possesses anti-tumoral activity by targeting multiple signaling transduction pathways. Several clinical trials employing AT-101 have been performed and some of them are still ongoing. Accordingly, we investigated the in vitro effects of AT-101 on cell proliferation, cell cycle regulation, pro-survival signaling pathways, apoptosis and autophagy of human (MM-B1, H-Meso-1, and MM-F1) and mouse (#40a) MM cell lines. In addition, we explored the in vivo anti-tumor activities of AT-101 in a mouse model, in which the transplantation of MM cells induces ascites in the peritoneal space. AT-101 inhibited in vitro MM cells survival in a dose- and time-dependent manner and triggered autophagy, but the process was then blocked and was coincident with apoptosis activation. To confirm the effect of AT-101 in inducing the apoptosis of MM cells, MM cells were simultaneously treated with AT-101 and with the caspase inhibitor, Z-VAD-FMK. Z-VAD-FMK was able to significantly reduce the number of cells in the subG1 phase compared to the treatment with AT-101 alone. This result corroborates the induction of cell death by apoptosis following treatment with AT-101. Indeed, Western blotting results showed that AT-101 increases Bax/Bcl-2 ratio, modulates p53 expression, activates caspase 9 and the cleavage of PARP-1. In addition, the treatment with AT-101 was able to: (a) decrease the ErbB2 protein expression; (b) increase the EGFR protein expression; (c) affect the phosphorylation of ERK1/2, p38 and AKT; (d) stimulate JNK1/2 and c-jun phosphorylation. Our in vivo results showed that the intraperitoneal administration of AT-101 increased the median survival of C57BL/6 mice intraperitoneally transplanted with #40a cells and reduced the risk of developing tumors. Our findings may have important implications for the design of MM therapies by employing AT-101 as an anticancer agent in combination with standard therapies.
RESUMEN
PURPOSE: Gossypol, a naturally occurring compound in cottonseeds, has anticancer effects against several tumor cell lines. It has been extensively studied in clinical trials and is well tolerated with a favorable safety profile. AT-101, a derivative of R (-)-gossypol, binds to Bcl-2 family proteins and induces apoptosis in vitro. Although transsphenoidal surgical excision of the pituitary corticotroph adenoma is the gold standard of care, it is not successful all the time. Medical therapy for Cushing's disease still remains a challenge for the clinicians. We aimed to investigate the cytotoxic and apoptotic effects of AT-101 in mouse pituitary corticotroph tumor AtT20 cells. METHODS: Cytotoxic effect of AT-101 was assessed by XTT cell viability assay. Apoptosis was shown by measuring DNA fragmentation and Caspase-3/7 activity. Changes in mRNA expressions of apoptosis-related genes were investigated by qPCR array after treatment with AT-101. ACTH was measured by ACTH-EIA Kit. RESULTS: AT-101 induced cytotoxicity and apoptosis in AtT20 cells. mRNA levels of pro-apoptotic genes such as TNFR-SF-10B, Bid, PYCARD, Caspase-8, Caspase-3, and Caspase-7 were induced by 2.0-, 1.5-, 1.7-, 1.5-, 1.6-, and 2-fold, respectively, in AtT20 cells by AT-101 treatment. Moreover, some of the anti-apoptotic genes such as BCL2L10, NAIP1, and PAK-7 were reduced by 2.1-, 2.3-, 4.0-fold, respectively, in AtT20 cells. AT-101 also decreased ACTH secretion significantly. CONCLUSION: AT-101 induces apoptosis in mouse pituitary corticotroph tumor cells.
Asunto(s)
Adenoma Hipofisario Secretor de ACTH/tratamiento farmacológico , Adenoma/tratamiento farmacológico , Hormona Adrenocorticotrópica/antagonistas & inhibidores , Apoptosis/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Gosipol/análogos & derivados , Neoplasias Hipofisarias/tratamiento farmacológico , Adenoma Hipofisario Secretor de ACTH/metabolismo , Adenoma Hipofisario Secretor de ACTH/patología , Adenoma/metabolismo , Adenoma/patología , Hormona Adrenocorticotrópica/metabolismo , Animales , Antineoplásicos Fitogénicos/farmacología , Proteínas Reguladoras de la Apoptosis/metabolismo , Gosipol/farmacología , Ratones , Neoplasias Hipofisarias/metabolismo , Neoplasias Hipofisarias/patología , Células Tumorales CultivadasRESUMEN
Malignant peripheral nerve sheath tumors (MPNSTs) are aggressive, Schwann cell-derived neoplasms of the peripheral nervous system that have recently been shown to possess an autocrine CXCL12/CXCR4 signaling loop that promotes tumor cell proliferation and survival. Importantly, the CXCL12/CXCR4 signaling axis is driven by availability of the CXCL12 ligand rather than CXCR4 receptor levels alone. Therefore, pharmacological reduction of CXCL12 expression could be a potential chemotherapeutic target for patients with MPNSTs or other pathologies wherein the CXCL12/CXCR4 signaling axis is active. AT101 is a well-established BCL-2 homology domain 3 (BH3) mimetic that we recently demonstrated functions as an iron chelator and thus acts as a hypoxia mimetic. In this study, we found that AT101 significantly reduces CXCL12 mRNA and secreted protein in established human MPNST cell lines in vitro. This effect was recapitulated by other BH3 mimetics [ABT-737 (ABT), obatoclax (OBX) and sabutoclax (SBX)] but not by desferrioxamine (DFO), an iron chelator and known hypoxia mimetic. These data suggest that CXCL12 reduction is a function of AT101's BH3 mimetic property rather than its iron chelation ability. Additionally, this study investigates a potential mechanism of BH3 mimetic-mediated CXCL12 suppression: liberation of a negative CXCL12 transcriptional regulator, poly (ADP-Ribose) polymerase I (PARP1) from its physical interaction with BCL-2. These data suggest that clinically available BH3 mimetics might prove therapeutically useful at least in part by virtue of their ability to suppress CXCL12 expression.
Asunto(s)
Antineoplásicos/farmacología , Compuestos de Bifenilo/farmacología , Gosipol/análogos & derivados , Imitación Molecular , Neurilemoma/tratamiento farmacológico , Nitrofenoles/farmacología , Pirroles/farmacología , Sulfonamidas/farmacología , Línea Celular Tumoral , Quimiocina CXCL12/genética , Quimiocina CXCL12/metabolismo , Regulación hacia Abajo , Regulación Neoplásica de la Expresión Génica , Gosipol/farmacología , Humanos , Indoles , Neurilemoma/genética , Neurilemoma/metabolismo , Neurilemoma/patología , Piperazinas/farmacología , Poli(ADP-Ribosa) Polimerasa-1/metabolismo , Unión Proteica , Dominios y Motivos de Interacción de Proteínas , Proteínas Proto-Oncogénicas c-bcl-2/metabolismoRESUMEN
BACKGROUND: Although epidermal growth factor receptor tyrosine kinase inhibitors (EGFR TKIs) have become the standard care of patients with advanced EGFR-mutant non-small cell lung cancer (NSCLC), development of acquired resistance is inevitable. A secondary mutation of threonine 790 (T790M) is associated with approximately half of the cases of acquired resistance. Strategies or agents to overcome this type of resistance are still limited. In this study, enhanced antitumor effect of AT-101, a-pan-Bcl-2 inhibitor, on gefitinib was explored in NSCLC with T790M mutation. METHODS: The effect of cotreatment with AT-101 and gefitinib on the viability of NSCLC cell lines harboring acquired T790M mutation was investigated using the MTT assay. The cellular apoptosis of NSCLC cells after cotreatment with AT-101 and gefitinib was assessed by FITC-annexin V/PI assay and Western blots analysis. The potential underlying mechanisms of the enhanced therapeutic effect for AT-101 was also studied using Western blots analysis. The in vivo anti-cancer efficacy of the combination with AT-101 and gefitinib was examined in a mouse xenograft model. RESULTS: In this study, we found that treatment with AT-101 in combination with gefitinib significantly inhibited cell proliferation, as well as promoted apoptosis of EGFR TKIs resistant lung cancer cells. The apoptotic effects of the use of AT-101 was related to the blocking of antiapoptotic protein: Bcl-2, Bcl-xl, and Mcl-1 and downregrulation of the molecules in EGFR pathway. The observed enhancements of tumor growth suppression in xenografts supported the reverse effect of AT-101 in NSCLC with T790M mutation, which has been found in in vitro studies before. CONCLUSIONS: AT-101 enhances gefitinib sensitivity in NSCLC with EGFR T790M mutations. The addition of AT-101 to gefitinib is a promising strategy to overcome EGFR TKIs resistance in NSCLC with EGFR T790M mutations.